The long-range goals are to (i) define the ontogeny and regulation of interstitial collagen formation in the amnion, (ii) explore the relationship between the regulation of amnion mesenchymal cell replication and the capacity for interstitial pro-collagen synthesis/processing in these unique cells, (iii) define selected collagen-related characteristics of the "dependent" portion of the fetal membranes; and (iv) explore the cause of increased risk for pre-term premature rupture of the fetal membranes in pregnancies of women who smoke cigarettes. It has been established that the interstitial collagens of the amnion zona compacta (the principal source of fetal membrane tensile strength) are synthesized exclusively in the widely dispersed mesenchymal cells of this avascular tissue. The greatest apparent capacity for interstitial collagen synthesis in amnion occurs early in pregnancy; beginning as early as 12-14 weeks gestation, the apparent capacity for pro-collagen synthesis/processing in amnion declines abruptly, perhaps in parallel with the decrease in the density of mesenchymal cells in this tissue. Studies are described to ascertain if the decline in the apparent capacity for interstitial collagen formation in amnion is related to a decreasing density of amnion mesenchymal or, rather, is the result of a decreased in the capacity for collagen formation in the mesenchymal cells per se. The final step in collagen formation involves the cross-linking of collagen fibrils which establish tensile strength and resistance to degradation by non-specific proteases. The initial reaction in collagen cross-linking is catalyzed by lysyl oxidase (LOX), a copper (Cu/2+)-dependent enzymes. The specific activity of LOX in amnion also declines strikingly after 12-14 weeks gestation. In approximately 25% of amnions examined during the third trimester, LOX enzyme activity is very low or undetectable, much lower than would be expected from the level of immunoreactive LOS protein in the same tissue. This finding of low LOX activity/per unit LOX protein is suggestive of an amnion mesenchymal cell-specific Cu/2+ deficiency. Studies are described to evaluate the hypothesis that cadmium in amniotic fluid (AF) (from maternal blood in women who smoke) acts upon amnion epithelial cells to induce the formation of metallothioneins (MT), which are metal binding proteins. Increased levels of MT in AF/amnion epithelial cells could sequester Cu?2+ and prevent its transfer to the mesenchymal cells, the site of LOX enzyme synthesis, creating a mesenchymal cell- specific Cu/2+ deficiency in pregnancies of women who, themselves, are not Cu/2+ deficient.